ABSTRACT Parkinson?s disease (PD) affects more than a million individuals in the U.S. with up to 60,000 new cases diagnosed each year. Currently, there are no treatments that can halt or reverse the course of PD; only palliative therapies, such as replacement strategies for missing neurotransmitters, exist. The pathobiology of PD is associated with the loss of dopaminergic (DA) neurons. Thus, the successful delivery of neurotrophic factors, in particular, glial cell line-derived neurotrophic factor (GDNF), that promote neuronal survival and reverse the progression of PD is of great importance. Regrettably, the blood brain barrier (BBB) remains a seemingly insurmountable obstacle to the routine use of systemically administered macromolecules - including GDNF1. To circumvent this problem, we propose using genetically modified peripheral blood monocytes (PBM) for systemic gene delivery to the brain. It is well established that specialized cells of the immune system, including monocytes, macrophages, and T cells, can easily penetrate the BBB and migrate rapidly to sites of brain inflammation and degeneration2,3. Our research has previously demonstrated that macrophages transfected ex vivo with therapeutic protein-encoding DNA plasmid (pDNA), can deliver therapeutic gene in intoxicated mice with acute brain inflammation, and in the transgenic mice, Parkin-Q311X(A). Mechanistic studies revealed that genetically modified macrophages release extracellular vesicles, exosomes, packed with protein-encoding genetic material, pDNA and mRNA, as well as a transcription factor involved in the encoded gene expression. Importantly, multiple lines of evidence for therapeutic efficacy were observed in PD mouse models, including decreased brain inflammation, significant neuroprotection, and improved locomotor functions. Planned studies include the evaluation of brain bioavailability for engineered PBM and GDNF gene transfer in Parkin-Q311X(A) mouse model (SA1). To enforce outcomes of the new formulation, PBM will be differentiated to a specific subset of ?alternatively activated? (M2) macrophages with regenerative functions. The mechanism of macrophage-mediated gene transfer including involvement cell-cell interactions and/or exosomes secreted by GDNF-transfected PBM will be elucidated (SA2). We will then assess the therapeutic potential of this novel product by measuring its anti-inflammatory and neuroprotective effects and, lastly, by extensive behavioral analysis in Parkin-Q311X(A) mice (SA3). To provide translational link, human induced pluripotent stem cells (iPSCs) with almost unlimited expandability will be tested. Furthermore, to obtain a universal cell-carrier, iPSCs with knockout MHC class II (MHC-II) receptor will be utilized. Our studies will provide fundamental insights into how PBM interact with brain cells and facilitate horizontal gene transfer upon neurodegeneration, potentially opening up other cell-based gene delivery systems to the CNS and beyond.